Fig. 2.
Fig. 2. Southern blots of 4 myeloma marrow samples with illegitimate recombinations. / The g denotes germline; L, legitimate recombination; *, illegitimate recombination; M, λHindIII molecular size marker, sizes indicated in kb. (A) Patient 1. IgAκ myeloma (1) μ to α isotype switch detected by matched 7.7-kb fragments inHindIII-5′Sμ and 3′Sα, and 8.0-kb fragments inSphI-5′Sμ and 3′Sα. Lanes 1 and 2 ofSphI-5′Sμ represent 2 hybridizations of separateSphI digests. (2) Illegitimate fragments detected in (i)HindIII-3′Sγ, 12 kb; absent in HindIII-3′Sα and 5′Sα, (ii) SphI-3′Sα and 5′Sα, 4.9 kb, suggesting a translocation upstream of 5′Sα or downstream of 3′Sα; absent inSphI-5′Sμ and 3′Sγ. The 5.2-kb band shown inBglII-3′Sα is also unmatched in BglII-5′ςμ or 5′Sμ, 5′Sα, and 3′Sγ (latter two not shown). Thus the presence of an illegitimate recombination in Sα has been confirmed by 2 restriction enzymes. (iii) BglII-5′ςα showed 9.4-kb and 2.0-kb fragments (the latter also detected by the 5′Sμ probe in close proximity to it), not present in Bgl II-3′Sα, 5′Sα, and 3′Sγ (latter two not shown). These recombinant bands are confirmed by 10 and 6.5 kb HindIII fragments detected by 5′ςμ. (NB: The 5′ςμ probe is located upstream of the HindIII site 5′ of the Sμ region, such that the 5′ςμ and 5′Sμ probes are located 5′ and 3′ of the HindIII site, respectively. Hence, the HindIII germline fragment detected by 5′ςμ does not contain the Sμ switch region.). (B) Patient 2. IgGκ myeloma (1) μ to γ isotype switch detected by 2.4-kb HindIII and 5.7-kbSphI fragments hybridized by 5′Sμ and 3′Sγ. Of the two recombinant BglII fragments (10.8 and 4.8 kb) hybridized by the 5′Sμ and 5′ςμ probes, the 10.8-kb fragment is likely to represent the legitimate switch, a matched fragment hybridized by 3′Sγ being obscured by the germline bands. (2) Illegitimate 6.4-kbHindIII fragment hybridized by 5′Sμ; absent inHindIII-3′Sγ and 3′Sα. This is confirmed by the 6.5-kbSphI fragment hybridized by 5′Sμ and the 4.8-kbBglII fragments hybridized by both 5′Sμ and 5′ςμ. Thus, the illegitimate recombination involving Sμ has been confirmed by 3 restriction enzymes. A nonspecific (NS) HindIII fragment that was hybridized by 3′Sγ was detected in both myeloma and germline control digests. (C) Patient 3. IgAκ myeloma (1) μ to α isotype switch detected by matched 6.6-kb HindIII fragments and 5.2-kb BglII fragments hybridized by 5′Sμ and 3′Sα. (2) Illegitimate 4.5-kb HindIII fragment hybridized by 5′Sμ but not by 3′Sα or 3′Sγ. This may be confirmed by the 9.6-kbBglII fragment detected by the 5′Sμ but not the 3′Sα probe, although a matching fragment hybridized by the 3′Sγ probe may be obscured by the BglII germline bands. (3) Illegitimate 3.5-kb HindIII fragment hybridized by 3′Sγ; not detected by the 5′Sμ and 3′Sα probes. (D) Patient 4. IgGκ myeloma (1) μ to γ isotype switch demonstrated by matched 7.8-kb SphI fragments hybridized by 5′Sμ and 3′Sγ. The 4.4-kbHindIII fragment hybridized by 5′Sμ probably represents the same legitimate switch, but a matching 3′Sγ fragment would be obscured by the germline fragment of similar size. (2) Illegitimate 7.4-kb HindIII fragment hybridized by 3′Sγ; absent in hybridization with 5′Sμ, 3′Sμ, 5′Sγ, 5′Sα, and 3′Sα. A nonspecific (NS) HindIII fragment that was hybridized by 3′Sγ was detected in both myeloma and germline control digests.

Southern blots of 4 myeloma marrow samples with illegitimate recombinations.

The g denotes germline; L, legitimate recombination; *, illegitimate recombination; M, λHindIII molecular size marker, sizes indicated in kb. (A) Patient 1. IgAκ myeloma (1) μ to α isotype switch detected by matched 7.7-kb fragments inHindIII-5′Sμ and 3′Sα, and 8.0-kb fragments inSphI-5′Sμ and 3′Sα. Lanes 1 and 2 ofSphI-5′Sμ represent 2 hybridizations of separateSphI digests. (2) Illegitimate fragments detected in (i)HindIII-3′Sγ, 12 kb; absent in HindIII-3′Sα and 5′Sα, (ii) SphI-3′Sα and 5′Sα, 4.9 kb, suggesting a translocation upstream of 5′Sα or downstream of 3′Sα; absent inSphI-5′Sμ and 3′Sγ. The 5.2-kb band shown inBglII-3′Sα is also unmatched in BglII-5′ςμ or 5′Sμ, 5′Sα, and 3′Sγ (latter two not shown). Thus the presence of an illegitimate recombination in Sα has been confirmed by 2 restriction enzymes. (iii) BglII-5′ςα showed 9.4-kb and 2.0-kb fragments (the latter also detected by the 5′Sμ probe in close proximity to it), not present in Bgl II-3′Sα, 5′Sα, and 3′Sγ (latter two not shown). These recombinant bands are confirmed by 10 and 6.5 kb HindIII fragments detected by 5′ςμ. (NB: The 5′ςμ probe is located upstream of the HindIII site 5′ of the Sμ region, such that the 5′ςμ and 5′Sμ probes are located 5′ and 3′ of the HindIII site, respectively. Hence, the HindIII germline fragment detected by 5′ςμ does not contain the Sμ switch region.). (B) Patient 2. IgGκ myeloma (1) μ to γ isotype switch detected by 2.4-kb HindIII and 5.7-kbSphI fragments hybridized by 5′Sμ and 3′Sγ. Of the two recombinant BglII fragments (10.8 and 4.8 kb) hybridized by the 5′Sμ and 5′ςμ probes, the 10.8-kb fragment is likely to represent the legitimate switch, a matched fragment hybridized by 3′Sγ being obscured by the germline bands. (2) Illegitimate 6.4-kbHindIII fragment hybridized by 5′Sμ; absent inHindIII-3′Sγ and 3′Sα. This is confirmed by the 6.5-kbSphI fragment hybridized by 5′Sμ and the 4.8-kbBglII fragments hybridized by both 5′Sμ and 5′ςμ. Thus, the illegitimate recombination involving Sμ has been confirmed by 3 restriction enzymes. A nonspecific (NS) HindIII fragment that was hybridized by 3′Sγ was detected in both myeloma and germline control digests. (C) Patient 3. IgAκ myeloma (1) μ to α isotype switch detected by matched 6.6-kb HindIII fragments and 5.2-kb BglII fragments hybridized by 5′Sμ and 3′Sα. (2) Illegitimate 4.5-kb HindIII fragment hybridized by 5′Sμ but not by 3′Sα or 3′Sγ. This may be confirmed by the 9.6-kbBglII fragment detected by the 5′Sμ but not the 3′Sα probe, although a matching fragment hybridized by the 3′Sγ probe may be obscured by the BglII germline bands. (3) Illegitimate 3.5-kb HindIII fragment hybridized by 3′Sγ; not detected by the 5′Sμ and 3′Sα probes. (D) Patient 4. IgGκ myeloma (1) μ to γ isotype switch demonstrated by matched 7.8-kb SphI fragments hybridized by 5′Sμ and 3′Sγ. The 4.4-kbHindIII fragment hybridized by 5′Sμ probably represents the same legitimate switch, but a matching 3′Sγ fragment would be obscured by the germline fragment of similar size. (2) Illegitimate 7.4-kb HindIII fragment hybridized by 3′Sγ; absent in hybridization with 5′Sμ, 3′Sμ, 5′Sγ, 5′Sα, and 3′Sα. A nonspecific (NS) HindIII fragment that was hybridized by 3′Sγ was detected in both myeloma and germline control digests.

Close Modal
This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal